未经证实:尽管金黄色葡萄球菌是生物膜相关感染的主要原因,对这些生物膜内的脂质组学分布知之甚少。这里,对金黄色葡萄球菌生物膜横截面进行脂质组学定位,以研究水平生物膜层之间的异质性.
未经评估:S.金黄色葡萄球菌生物膜静态生长,包埋在羧甲基纤维素/明胶的混合物中,并准备用于下游基质辅助激光解吸/电离成像质谱(MALDIIMS)。在质量分析之前还应用捕获的离子迁移谱(TIMS)。
未经授权:TIMS的实施导致检测到的脂质种类数量增加了3倍。用甲酸铵(150mM)洗涤生物膜样品使一些细菌脂质的信号强度增加了十倍,对生物膜结构的破坏最小。MALDITIMS显示,大多数脂质主要位于单个生物膜层,和来自相同脂质类别的物种,如心磷脂CL(57:0)-CL(66:0)显示出截然不同的定位,层之间表现出1.5至6.3倍的强度差异(n=3,p<0.03)。在厌氧生长的生物膜内没有观察到水平层,脂质分布均匀。
UNASSIGNED:通过MALDITIMS对金黄色葡萄球菌生物膜横截面的高空间分辨率分析揭示了水平金黄色葡萄球菌生物膜层之间明显的脂质组学异质性,表明每个层在分子上是不同的。最后,这个工作流程揭示了在厌氧条件下生长的生物膜中不存在层,可能表明氧气有助于在有氧条件下观察到的异质性。该工作流程在研究对抗菌药物的空间局部分子反应的未来应用可以提供新的治疗策略。
UNASSIGNED: Although Staphylococcus aureus is the leading cause of biofilm-related infections, the lipidomic distributions within these biofilms is poorly understood. Here, lipidomic mapping of S. aureus biofilm cross-sections was performed to investigate heterogeneity between horizontal biofilm layers.
UNASSIGNED: S. aureus biofilms were grown statically, embedded in a mixture of carboxymethylcellulose/gelatin, and prepared for downstream matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS). Trapped ion mobility spectrometry (TIMS) was also applied prior to mass analysis.
UNASSIGNED: Implementation of TIMS led to a ∼ threefold increase in the number of lipid species detected. Washing biofilm samples with ammonium formate (150 mM) increased signal intensity for some bacterial lipids by as much as tenfold, with minimal disruption of the biofilm structure. MALDI TIMS IMS revealed that most lipids localize primarily to a single biofilm layer, and species from the same lipid class such as cardiolipins CL(57:0) - CL(66:0) display starkly different localizations, exhibiting between 1.5 and 6.3-fold intensity differences between layers (n = 3, p < 0.03). No horizontal layers were observed within biofilms grown anaerobically, and lipids were distributed homogenously.
UNASSIGNED: High spatial resolution analysis of S. aureus biofilm cross-sections by MALDI TIMS IMS revealed stark lipidomic heterogeneity between horizontal S. aureus biofilm layers demonstrating that each layer was molecularly distinct. Finally, this workflow uncovered an absence of layers in biofilms grown under anaerobic conditions, possibly indicating that oxygen contributes to the observed heterogeneity under aerobic conditions. Future applications of this workflow to study spatially localized molecular responses to antimicrobials could provide new therapeutic strategies.